Grace Catalog 2008

hplc applications
Purification of Trityl-On Oligonucleotides
Proteins & Peptides/Reversed-Phase
Human Growth Hormone
Human Growth Hormone
3
1. di-Desamido HGH
1. di-desamido HGH
2. mono-Desamido 2. mono-desamido HGH HGH
3. Human Growth 3. human Hormone growth (HGH) hormone (HGH)
Purification of Trityl-On Oligonucleotides
36-mer
1
2
0 7 14 21 28 35 42 Min.
Column: Vydac ® C4, 5µm, 250 x 4.6mm (Part No. 214TP54)
Mobile Column: Phase: VYDAC Isopropanol:0.01M 214ATP54 (C4, 5 µm, Tris 4.6 mm buffer, i.d. pH7.5 x 250 mm) (29:71)
Flow Eluent: Rate: 29% isopropanol/71% 1.0mL/min 0.01 M Tris buffer, pH 7.5
Detector: UV
Separation of Protein / Peptide Mixture
on Vydac 219TP Diphenyl Column
Separation of Protein / Peptide Mixture on
Vydac ® 219TP Diphenyl Column
1
3
2
4
2
5
2 6
7
8
1. Horse Cytochrome C
Tryptic Digest Fragments
2. Whale Myoglobin CNBr
Digest Fragments
3. Ribonuclease
4. Cytochrome
5. Lysozyme
6. BSA
7. Myoglobin
8. Ovalbumin
0 5 10 15 20 Min.
Column: Vydac ® C4, 5µm, 250 x 10mm (Part No. 214TP510)
Mobile Phase: A: 0.1M Triethylammonium Acetate, pH7.0 B: Acetonitrile
Gradient: Column: VYDAC Time: 214TP510 0 14(C4, 5 µm, 10 mm i.d. x 250 mm)
Gradient: 16
B%:
to 23%
16
ACN
23
in 0.1 M triethylammonium acetate,
pH 7.0, over 14 minutes
Flow Comparison Rate: of 5.0mL/min
Polypeptide Separations on
Detector: C18, C4 and Diphenyl UV Reversed-Phase Columns
218TP
(C18)
214TP
(C4)
219TP
(diphenyl)
Comparison of Polypeptide Separations on
C18, C4 and Diphenyl Reversed-Phase Columns
1
1
1
2
2
2
1. H
2. TH6
life science
1. H
2. TH6
0 10 20 30 40 50 60 70 Min.
Column: Vydac ® Diphenyl, 5µm, 250 x 4.6mm
Columns: VYDAC (Part 219TP54 No. (diphenyl, 219TP54) 5 µm, 4.6 mm i.d. x 250 mm)
Gradient: Mobile Phase: 0-70% A: ACN 0.1% with Trifluoroacetic 0.1% TFA in 67.5 Acid min
Sample: 1. horse B: cytochrome Acetonitrilec tryptic digest fragments;
Gradient: 2. whale Time: myoglobin 0 CNBr 67.5 digest fragments;
3. ribonuclease; 4. cytochrome c; 5. lysozyme; 6. BSA;
B%: 0 70
IgG 7. Purification myoglobin; from and 8. Bovine ovalbumin Serum
Flow Rate: 1.0mL/min
Detector: UV
IgG Purification from Bovine Serum
2
1. Serum Load Passthrough
2. IgG 1. Serum Load Passthrough
2. IgG
0 5 10 15 20 25 30 Min.
Column: Vydac ® C18, C4 and Diphenyl, all 5µm, 150 x 4.6mm
(Part Nos. 218TP5415, 214TP5415, 219TP5415)
Columns: Mobile Phase: VYDAC A: 218TP5415 0.1% Trifluoroacetic (C18), 214TP5415 Acid (C4) and
219TP5415 B: Acetonitrile (diphenyl), all 5 µm, 4.6 mm i.d. x 150 mm
Gradient: 10-90% ACN with 0.1% TFA over 30 min
Gradient: Time: 0 30
Sample: 18-residue helical peptide (1) and six-helix templateassembled
B%: synthetic 10 90 protein (2)
Sample: 18-Residue Helical Peptide (1) and
Six-helix Template- assembled Synthetic Protein (2)
Flow Rate: 1.0mL/min
Detector: UV
Analysis of a Lipid Peptide
Analysis of a Lipid Peptide
1
0 5 10 15 20 25 30 35 40 45 Min.
0 10 20 30 40 50 60 Min.
Column: Vydac ® Venture ® A RAVE Recombinant Protein A,
Column: Vydac ® Diphenyl, 5µm, 250 x 4.6mm
15-20µm, 50 x 4.6mm (Part No. 1520450)
(Part No. 219TP54)
Mobile Column: Phase: VENTURE A: 20mM A RAVE1520450
Phosphate, pH7.4, 150mM Sodium Chloride Mobile Column: Phase: VYDAC A: 0.1% 219TP54 Trifluoroacetic (diphenyl, 5 µm, Acid 4.6 mm i.d. x 250 mm)
(Recombinant Protein A, 15-20 µm, 4.6 mm i.d. x 50 mm)
B: 0.1 M Sodium Citrate, pH3.2
Flow: 1.5 mL/min
B: n-Propanol:Acetonitrile:0.1% Trifluoroacetic Acid (50:50)
Flow:
1.38 mL/min (linear velocity = 500 cm/hr)
Temp: 45 °C
Gradient: Equilibrate with A for 3 min, load sample for 9 min,
Gradient: Time: 0 45
Eluent: A = 20 mM phosphate, pH 7.4, 150 mM NaCl
Eluent: A = 0.1% TFA (v/v) in water
B = 0.1 wash M sodium with A citrate, for 12 pH min, 3.2elute with B for 3 min,
B = 50:50 %B: n-propanol:ACN, 25 70 0.1% TFA (v/v)
Gradient: Equilibrate re-equilibrate with A for with 3 min, A for load 6 sample min for 9 min, wash
Flow Gradient: Rate: 25 to 1.5mL/min
70% B over 45 minutes
Flow Rate: with 1.38mL/min A for 12 min, elute (linear with velocity B for 3 = min, 500cm/hr) re-equilibrate
Detector: UV
Detector: with UV A for at 6280nm
min
Detection: Absorbance, 280 nm
more
Sample:
applications
Commercial bovine serum diluted 1:20 with A and filtered
View our complete through searchable a 0.45 chromatogram µm filter. Injection database volume at www.discoverysciences.com/chromdb/
= 12.4 mL.
ww.discoverysciences.com
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